Women are more susceptible than men to several aspects of drug addiction, including relapse due to stressful events. Addictive mechanisms require associative memory processes that critically involve hippocampal circuits, including the opioid system. Our light and electron microscopic (EM) studies have revealed notable sex and estrous cycle differences in the hippocampal opioid system: enkephalins, mu opioid receptors (MORs) and delta opioid receptors (DORs) are subcellularly positioned in neurons in the mossy fiber circuitry to enhance excitability and learning processes in females with elevated estrogen states. Moreover, unlike males, females in high estrogen states exhibit a long-term potentiation (LTP) evoked by low frequency stimulation of the mossy fibers that is regulated by DORs. Progress during the last grant period showed that chronic immobilization stress (CIS) in males, which results in CA3 neuron dendritic retraction and parvalbumin interneuron loss, essentially shuts off the hippocampal opioid system. Conversely, CIS in females, regardless of estrogen state, primes the opioid system for greater excitation of CA3 pyramidal cells. Specifically, afte CIS mossy fiber enkephalin levels are elevated and the distribution of MORs and DORs in hippocampal neurons resembles that seen in unstressed females at high estrogen states. Moreover, our EM studies revealed in females, but not males, the involvement of a second mechanism for enhancing hippocampal excitation following CIS: DORs traffic to the near plasmalemma of hilar GABAergic interneurons that are known to project to granule cells dendrites where they converge with entorhinal afferents. Notably one-hour after a single injection of oxycodone (3mg/kg, i.p.) the DORs in these GABAergic interneurons have moved to the plasmalemma where additional exposure to ligand would disinhibit these neurons and thus could promote LTP at entorhinal-granule cell synapses. To date, however, no studies have explored the mechanisms by which chronic stress-induced changes in the opioid system in females could advance associative memory processes important for drug addiction, particularly to oxycodone. Thus, this renewal application proposes to test the central hypothesis that the alterations in the hippocampal opioid system induced by chronic stress predispose females to enhanced sensitivity to oxycodone and promote drug-related associative learning. Aim 1 will first compare the effects of CIS on the acquisition of oxycodone-induced conditioned place preference (CPP) in females and males. Second, MORs and DORs will be selectively antagonized at critical hippocampal sites, to determine their role in oxycodone CPP in unstressed and CIS female and male rats. Aim 2 will determine how oxycodone CPP alters specific molecular (as assessed by targeted PCR gene array) and anatomic (as assessed by light and EM immunocytochemistry and in situ hybridization) profile of the hippocampal mossy fiber opioid system in female and male rats under basal conditions and after CIS.